Method for the spectroscopic examination of a biological tissue

ABSTRACT

In a method for spectroscopic examination of a biological tissue the tissue is irradiated with light having an intensity l 0  (λ) with the wavelength λ being varied, a transmission or reflection spectrum l(λ) is obtained by measuring the intensity of the radiation transmitted or reflected by the tissue dependent on the wavelength λ, and an approximate description of the measured transmission or reflection spectrum l(λ) or of the spectrum l(λ)/l 0  (λ) normalized to the incident radiation intensity, or of the quantity log {l(λ)/l 0  (λ)} with an analytical function is made. The analytical function has a Beer-Lambert dependency on a first parameter which describes the absorption properties of the tissue and has a second parameter which is the average path length L(λ) of the photons in the tissue, representing a slice thickness. The first parameter is dependent on concentrations c i  of a number i of selected tissue components which are employed as fit parameters, as well as being dependent on specific absorption coefficients α i  (λ) according to the relationship ##EQU1## The identified concentrations c 1  are then compared to reference values.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention is directed to a method for spectroscopicexamination of biological tissue.

2. Description of the Prior Art

The diagnosis of mammary carcinoma is currently mainly based on theimaging method of X-ray mammography. Portions of the public and of themedical community, however, are increasingly critical of thisexamination method since damage to the transirradiated tissue cannot beprecluded with certainty. Additionally utilized, further, is theextremely complicated nuclear magnetic resonance tomography as well as,to a slight extent, ultrasound measuring methods and infraredthermography.

Light-tomographic methods are being clinically tested wherein to tissueto be examined is illuminated with visible or, respectively, IR lightand the reflected or transmitted radiation is detected (see, forexample, G. Muller et al. (Eds.), Medical Optical Tomography. FunctionalImaging and Monitoring, The international Institute for OpticalEnineering sic!, Bellingham, Wash., 1993, SPIE Vol. IS11; B. Chance, R.R. Alfano, A. Katzir (Eds.) Photon Migration and Imaging in Random Mediaand Tissues, The International Institute for Optical Engineering,Bellingham, Wash. 1993, SPIE Vol. 1988; G A Navarro, A. E. Profio; Med.Phys. Vol. 15(1988); S. 181-187. Since the measured intensities aredependent on the optical properties of the respectively transirradiatedregion, one hopes to be able to distinguish tissue types and identifyand localize physiological or, respectively, pathological modificationsin the tissue. Possible applications of light-tomography extend from thedetection of mammary carcinoma via the recognition of Alzheimer'sdisease up to the registration of the oxygenation of the brain and theextremities.

SUMMARY OF THE INVENTION

An object of the invention is to create an optical method with whichphysiological and pathological modifications in a biological tissue,particularly in the human body, can be detected in vivo. The methodshould not produce any damage to the tissue and should be comparativelysimple to implement without great apparatus outlay.

The above object is achieved in a method for spectroscopic examinationof a biological tissue wherein the tissue is irradiated with lighthaving an intensity l₀ (λ) with the wavelength λ being varied, atransmission or reflection spectrum l(λ) is obtained by measuring theintensity of the radiation transmitted or reflected by the tissuedependent on the wavelength λ, and an approximate description of themeasured transmission or reflection spectrum l(λ) or of the spectruml(λ)/l₀ (λ) normalized to the incident radiation intensity, or of thequantity log {l(λ)/l₀ (λ)} with an analytical function is made. Theanalytical function has a Beer-Lambert dependency on a first parameterwhich describes the absorption properties of the tissue and has a secondparameter which is the average path length L(λ) of the photons in thetissue, representing a slice thickness. The first parameter is dependenton concentrations c_(i) of a number i of selected tissue componentswhich are employed as fit parameters, as well as being dependent onspecific absorption coefficients α_(i) (λ) according to the relationship##EQU2## The identified concentrations c₁ are then compared to referencevalues.

The method is preferably employed in the area of medical diagnostics.Due to its enhanced sensitivity or, respectively, selectivity comparedto what is referred to as diaphanography (see, for example, /3/),regions with pathological tissue modifications can be distinguishedbetter from surrounding, healthy tissue and carcinoma in the femalebreast can be more exactly localized.

DESCRIPTION OF THE DRAWINGS

FIGS. 1 and 2 show the absorption coefficients of hemoglobin (H_(b)),oxyhemoglobin (H_(b) O), water and fat (vegetable oil) dependent onwavelength.

FIG. 3 shows the reduced scatter coefficient of breast tissue (in vivo)dependent on the wavelength.

FIG. 4 is a schematic illustration of an apparatus for registering atransmission spectrum according to the method of the invention.

FIGS. 5 through 8 illustrate measured in vivo transmission spectra of afemale breast obtained in accordance with the principles of the presentinvention.

DESCRIPTION OF THE PREFERRED EMBODIMENT

The invention strives to describe the transmission spectra of the femalebreast measured in vivo by an analytical function l(λ), whereby thefollowing, highly simplified model assumptions form the basis:

1) the breast tissue is composed of a uniform mixture of the absorberswater, fat, Hb and HbO;

2) the sum of the volume parts of water and fat amounts to 100%;

3) the volume parts of Hb and HbO are left out of consideration (2.3mMol per liter of blood).

In an exclusively absorbent medium, the attenuation of a light rayexhibiting the initial intensity l₀ is described by the Beer-Lambert law

    l=l.sub.0 ·exp {-μ.sub.a ·d}          (1)

whereby d references the thickness of the transirradiated slice andμ_(a) references the absorption coefficient of the medium. When themedium is composed of a plurality of absorbers, then the absorptioncoefficient μ_(a) is calculated from the sum of the products of theabsorber concentrations c_(i) and the specific absorption coefficientsα_(i) as ##EQU3##

In the case of a tissue composed of the above-recited components, μ_(a)is thus established by

    μ.sub.a =C.sub.water ·α.sub.water +C.sub.fat ·α.sub.fat +C.sub.Hb ·α.sub.Hb +C.sub.HbO ·α.sub.HbO                                 (3)

Equation (1), however, is not valid for highly scattering media sincethe path lengths covered by the individual photons in the medium are notknown here. Patterson et al. Appl. Optics, Vol. 28 (1988), pp 2331-2336,however, have solved the diffusion equation for a plane-parallelgeometry and calculated the average path length L(λ) of the photons in ascattering medium as ##EQU4## whereby μ_(s) ' references thewavelength-dependent, reduced scatter coefficient of the medium. Whenthe thickness d in the Beer-Lambert law is then replaced by thewavelength-dependent average path length L(λ) of the photons andwavelength-independent correction factors x₁, x₂ are introduced, thenthe transmitted intensity l(λ) can be written as

    l(λ)=l.sub.0 (λ)·x.sub.1 ·exp{-μ.sub.s (λ)·L(λ)·x.sub.2 }        (5)

The correction factor x₁ takes the measurement geometry and the multiplyscattered, unabsorbed, undetected photons into consideration.Measurements errors of the slice thickness d and imprecisions of thereduced scatter coefficient μ_(s) ' are intended to be compensated bythe factor x₂ . Given knowledge of the absorption coefficients α_(i),the analytical function according to Equation (5) can be matched tomeasured in vivo spectra by variation of the concentration c_(i) of thefour tissue components and of the two correction factors x₁, x₂.

The Absorption Coefficients of the Tissue Components Blood, Water andFat

For the in vivo measurement of the transmission spectrum of thickbiological tissue (d≧3 cm), the principal absorbers blood, water and fatdefine the diagnostic window. It begins in the short-wave range at aboutλ=600 nm (blood) and ends in the long-wave range at about λ=1.4 mm(water). Due to the detector system (Si photodiode) employed, however,only measurements in the wavelength range between λ≧650 nm and λ≦1.1 μmwere carried out.

FIG. 1 shows the specific absorption coefficients of the blood pigmentshemoglobin Hb and oxyhemoglobin HbO in the wavelength range betweenλ=650 nm and λ=1000 nm. One can clearly see the absorption maximum of Hbat λ=760 nm. The absorption coefficients of Hb and HbO are the same sizeat the isobestic point (λ=805 nm).

The absorption coefficients respectively calculated from the measuredtransmission characteristic of water and fat (vegetable oil) are shownin FIG. 2. The O--H resonance of the water molecule has a highlyabsorbent effect at λ=975 nm. Weaker harmonics of this resonance areobserved at λ=840 nm and λ=755 nm. Compared thereto, fat or,respectively, the vegetable oil serving as substitute absorbs especiallystrongly in the region of the C--H resonance (λ=930 nm). The absorptionis considerably weaker in the region of λ=760 and λ≈830 nm, where FIG. 2exhibits only very flat structures.

The Reduced Scatter Coefficient of the Breast Tissue

The calculation of the dependency of the reduced scatter coefficient onthe wavelength is only possible given knowledge of the size andcomposition of the scatter centers. Since corresponding information forthe breast tissue are lacking, our own measurements were carried out atvarious wavelengths. FIG. 3 shows the results of these measurements. Theinterpolation lines based on the data and the in vivo or, respectively,in vitro measured values of other authors are also shown. The reducedscatter coefficient is only weakly dependent on the wavelength and lieson the order of magnitude of 1 mm⁻¹.

Experimental Structure

FIG. 4 shows the schematic structure of an apparatus for theregistration of an in vivo transmission spectrum. This is therebyessentially a matter of a commercially available spectroradiometer(Merlin, ORIEL Company) whose components were adapted to the newmeasurement technology. A 100 W halogen lamp 1 that has a uniform andhigh spectral irradiance in the measurement range (500 nm≦λ≦1100 nm)serves as white light source. Controlled by the computer 2, the gridmonochromator 3 (600 lines/mm; linear reciprocal dispersion 12.8 nm/mmat λ=750 nm) resolves the light emitted by the halogen lamp 1 into itsspectral components. The selected component subsequently passes throughthe edge filter 4 arranged following the monochromator 3, so that theoptics 5 couples only the radiation diffracted in the first order intothe optical fiber cable 7 connected to the fiber head 6 (diameter: 5mm). Since the radiant intensity at the fiber head amounts toapproximately 1 mW (λ=800 nm, width of the monochromator slit s=1 mm),the power density on the skin--at approximately 5 mW/cm² --lies farbelow the allowable limit value. The radiation penetrates into thebreast 9 lying on the transparent plate 8, is absorbed and scattered toa greater or lesser extent according to the optical properties of thetransilluminated tissue and in turn emerges at the side lying oppositethe optical fiber head 6. An approximately 2 m long optical fiber cable10 acquires the transmitted radiation and conducts it to the Siphotodiode.

The beam interrupter 12 (propeller wheel) arranged in the beam pathbetween the halogen lamp 1 and the monochromator 3 has the job ofmodulating the intensity of the light entering into the monochromatorand, thus, the intensity of the radiation illuminating the breast tissue9 with a frequency of, for example, f=25 to 30 Hz prescribed by thedrive unit 13.

As a result, the output signal of the Si photodiode 11 also has acomponent exhibiting the modulation frequency f whose amplitude isidentified in the lock-in amplifier 14 and read into the computer 2. Theforked light barrier composed of a light-emitting diode 15 and aphotodiode 16 generates the reference signal adjacent at a second inputof the lock-in amplifier 14.

Experimental Results

The in vivo transmission spectra described briefly below were registeredwith the apparatus shown in FIG. 4. Approximately 150 s are required forthe registration of the respectively 50 measured values per spectrum(step width Δλ=10 nm). During the measurement, the widths of the inputand output gap of the monochromator 3 were respectively set to 1 mm.This corresponds to a bandwidth of approximately 13 nm, which assures anadequately high spectral resolution. The quantity

    log{l(λ)/l.sub.0 (λ)}                        (6)

referred to below as transmittance was respectively evaluated. It isshown in FIGS. 5 through 8 together with the corresponding simulationcurves dependent on the wavelength. A table with the corresponding fitparameters is respectively allocated to the individual spectra. Asfurther parameters, the tables or, respectively, Figures potentiallyalso contain the age of the test subject, particulars about the locationof the measurement on the breast and the thickness of thetransilluminated tissue.

Despite the differences in the height of the respectively measuredtransmittance, all in vivo spectra agree in terms of the followingfeatures:

1) below λ=600 nm (blood absorption), the transmittance drops by anumber of orders of magnitude;

2) a transmission minimum is observed at approximately λ=760 nm overlapof the Hb signal and the weaker fat and water signals);

3) more or less pronounced minimums show up at λ=975 nm (fat) and λ=930nm (water);

4) the transmittance decreases within the entire wavelength range withincreasing blood content of the tissue.

FIGS. 5 and 6 show representative in vivo transmission spectra of thefemale breast, whereby FIG. 5 is directed to the measurementsimplemented at younger persons (40 years old and younger) and FIG. 6 isdirected to the measurements implemented at older persons (60 years oldand older). It is not difficult to see with reference to the fitparameters recited in Table I that younger and older breast tissuediffer noticeably from one another with respect to the fat, water andblood content.

                  TABLE I                                                         ______________________________________                                        Fit Parameter:                                                                         younger persons older persons                                        ______________________________________                                        Age      39      37      33    85    80    64                                 Water  %!                                                                              33      65      70    17    11    12                                 Fat  %!  67      35      30    83    89    88                                 Hb  1/min!                                                                             0.0012  0.0017  0.002 0.0005                                                                              0.0002                                                                              0.0005                             HbO  1/mm!                                                                             0.0042  0.0042  0.0041                                                                              0.003 0.0014                                                                              0.0018                             x.sub.1  0.0002  0.0005  0.0007                                                                              0.0004                                                                              0.0006                                                                              0.0005                             x.sub.2  2.1     2.1     2.1   1.9   1.65  1.85                               ______________________________________                                    

FIG. 7 shows the influence of the measurement position on the in vivotransmission spectrum. Only the spectra measured at the positions "A"and "D" are shown. With the parameters recited in Table II, one againsucceeds in describing the spectra very well by the function l(λ)recited in Equation (5).

                  TABLE II                                                        ______________________________________                                        Fit Parameter:                                                                Position ref      A        B      C     D                                     ______________________________________                                        Water  %!                                                                              24       24       20     16    12                                    Fat  %!  76       76       80     84    88                                    Hb  1/mm!                                                                              0.0005   0.0005   0.0005 0.0005                                                                              0.0003                                HbO  1/mm!                                                                             0.0023   0.0024   0.0024 0.0025                                                                              0.0025                                X1       0.0006   0.0006   0.0006 0.0006                                                                              0.0006                                X2       1.4      1.53     1.6    1.6   1.6                                   ______________________________________                                    

As FIG. 8 shows, the transmission spectrum (black squares) measured atan unhealthy breast (invasive duct carcinoma) differs clearly,particularly in the wavelength range between λ=900 nm and λ=1000 nm,from the reference spectrum registered at the corresponding location ofthe healthy breast (unfilled squares). This can be essentiallyattributed to the modified water and fat content in the region of thecarcinoma compared to the healthy breast. Moreover, the transmittancemeasured at the unhealthy breast is clearly lower than in the referencespectrum at all wavelengths.

The invention, of course, is not limited to the described exemplaryembodiments. Thus, it is possible without difficulty to also registerreflection spectra and to again describe these by Equation (5). Thequantity L(λ) is then established by ##EQU5## r: source-detector spacing(see Farell et al., Med. Phys. 19(4)(1992), pp. 879-889).

We claim as our invention:
 1. Method for the spectroscopic examinationof a biological tissue comprising the steps of:(a) irradiating a tissueregion with non-ionizing radiation having an intensity l₀ (λ) and awavelength λ while varying said wavelength; (b) registering a spectruml(λ) from an element selected from the group consisting of atransmission spectrum and a reflection spectrum by an element selectedfrom the group consisting of measuring the intensity of the radiationtransmitted by the tissue and radiation reflected by the tissuedependent on the wavelength λ; (c) employing an analytical function toapproximately describe a quantity from an element selected from thegroup consisting of said spectrum l(λ), said spectrum l(λ)/l₀ (λ) normedto the incident radiation intensity, and log {l(λ)/l₀ (λ)}, saidanalytical function having a Beer-Lambert dependency on a firstparameter describing the absorption properties of the tissue and asecond parameter representing a slice thickness, the first parameterbeing dependent on identified concentrations c_(i) of a plurality i ofselected tissue components employed as fit parameters as well as onspecific absorption coefficients α_(i) (λ) according to the relationship##EQU6## with an average path length L(λ) of the photons in the tissuecomprising the second parameter; and (d) comparing the identifiedconcentration c_(i) to reference values.
 2. Method according to claim 1,characterized in that the analytical function is established by theequation

    l(λ)=l.sub.0 (λ)·x.sub.1 ·exp{-μ.sub.s (λ)·L(λ)·x.sub.2 }

whereby x₁, x₂ reference wavelength-independent correction factors. 3.Method according to claim 1 wherein the average path length L(λ) of thephotons in the tissue is dependent on a thickness d of thetransilluminated tissue, a reduced scatter coefficient μ_(s) '(λ) and onthe absorption coefficient μ_(a) of the tissue according to therelationship ##EQU7##
 4. Method according to claim 1 wherein the tissueis approximated as a uniform mixture of water, fat, hemoglobin andoxyhemoglobin, whereby volume parts of water and fat supplement to 100%.5. Method according to claim 1 comprising modulating the intensity ofthe incident radiation at a modulation frequency and registering anamplitude of an output signal of a detector system exhibiting themodulation frequency dependent on the wavelength.